Chiang Mai J. Sci.

2006; 33(1) : 123-128

www.science.cmu.ac.th/journal-science/josci.html
Contributed Paper

High Performance Thin Layer Chromatographic

Method for the Determination of Diclofenac Sodium
in Pharmaceutical Formulations
Wisanu Thongchai, Boonsom Liawruangrath*, Chalermporn Thongpoon
and Theeraphan Machan
Department of Pharmaceutical Science, Faculty of Pharmacy, Chiang Mai University, Chiang Mai 50200, Thailand.
* Author for Correspondence, e-mail: boonsom@pharmacy.cmu.ac.th
Received : 14 March 2005
Accepted : 24 June 2005

ABSTRACT
A high performance thin layer chromatographic method for the determination of
diclofenac sodium in pharmaceutical formulations was developed. The drug was extracted
from the sample (emulgel) then various aliquots of this solution were spotted automatically by
means of Camag Linomat IV (Switzerland) on a silica gel 60 F254 aluminium plate, using a
mixture of toluene : ethyl acetate : glacial acetic acid (60:40:1, v/v/v) as mobile phase. The
spot areas were quantified by densitometry at 282 nm. Linear calibration curve was obtained
over the range 5-80 g.mL-1 (r2 = 0.9993). The method was applied to the determination of
diclofenac sodium in Diclogel, Voltaren emulgel and Dosanac emulsiongel with the average
percentage recoveries of 104.211.59, 112.411.93 and 101.274.59, respectively. The standard
deviation of diclofenac sodium in Dosanac emulsiongel was higher than those obtained
from Diclogel and Voltaren emulgel, probably owing to the matrices present in the sample.
But the recoveries of the added diclofenac sodium in the samples are quite good. The average
percentage labelled amount of diclofenac sodium in Diclogel, Voltaren emulgel and Dosanac
emulsiongel were 94.610.06, 97.870.11 and 94.810.03, respectively. They are not exceed
the percentage labelled amount claimed, (not less than 90% and not more than 110%, USP 28)
[1]. The proposed method is simple, rapid, sensitive, reproducible and accurate. It also consumed
less reagents compared with the HPLC method. Therefore this method is suitable for routine
analysis of this drug in raw materials and formulations.
Keywords: diclofenac sodium, high performance thin layer chromatography, pharmaceutical
formulations.
1. I NTRODUCTION
Diclofenac sodium [Sodium (o- {(2,6dichlorophenyl) amino} phenyl) acetate]
(Figure 1) is a synthetic nonsteroidal antiinflammatory Drug (NSAID), has been
proven as a safe and efficacious drug in the
treatment of a variety of inflammatory and
rheumatoid disorders [2]. The pharmacological effects of this drug are thought to be related
to the inhibition of the conversion of arachi-

O
Cl

H
N

ONa

Cl

Figure 1. Structure of diclofenac sodium.

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donic acid to prostaglandins, which are the

mediators of the inflammatory process [3].
It is employed mainly in oral formulations,
and the some extent, also for intramuscular
injection and topical formulation [4]. For
topical formulations, one of the most wildly
used is Voltaren emulgel, Diclogel and
Dosanac emulsiongel. The advantage for this
type of dosage form found to be effective
for the treatment of local inflammatory [4].
However, the disadvantage was also observed
due to careless of the patients which may wipe
the drug out shortly after applied to the skin.
For the analysis of diclofenac, several
spectrophotometric methods have been
reported. Bucci et al. [5] described a UV ( =
276 nm) and UV first derivative ( = 240340 nm) spectrophotometric methods for the
determination of diclofenac sodium or
diethylammonium in tablets, suppositories and
gel with the limit of detection (LOD) of 4
mole.L -1 . Sena et al. [6] determined
diclofenac in pharmaceutical formulations
containing vitamin B by using UV spectrophotometry and partial least squares
regression. Bottello et al. [7] presented a
sensitive spectrophotometric method for
determining diclofenac sodium using methylene blue as colorimetric reagent in the pH
range 9.2-9.4 to form a chlorofor mextractable blue ion-association complex
which gives maximum absorption at 653 nm.
Agrawal et al. [8] reported two colorimetric
methods for determining diclofenac sodium
in tablets. The first method is based on the
reaction of diclofenac with ferric chloride and
2, 2- bipyridine to form a complex having
maximum absorption at 520 nm. The second
method is based on the reaction of diclofenac
with methylene blue in phosphate buffer pH
6.8 forming a complex which gives maximum
absorption at 640 nm. Damiani et al. [9]
reported a spectrofluorometric method for
quantitative analysis of diclofenac in 0.01 N
HCl and measured the fluorescence intensity
at em = 362 nm with the ex = 287 nm. Shafiee
et al. [10] determined diclofenac sodium in
injectable solution by gas chromatography

Chiang Mai J. Sci. 2006; 33(1)

(GC) and high performance liquid chromatography (HPLC). The ester was extracted and
subjected to gas liquid chromatography with
flame ionization detector, 5% SE-30/chrom
W-HP (80100 mesh) was used as a column
in GC. For reversed phase HPLC, MeOH :
H2O (55:45, v/v) was used as mobile phase.
The separation was performed on a bondapak phenyl column using UV detector
at 274 nm. Gonzalez et al. [11] determined
diclofenac sodium in the presence of cyanocobalamin and betamethasone in tablets by high
performance liquid chromatography using RP
18 column and acetonitrile : water (40:60, v/
v) (pH 3.45) as mobile phase with UV detection at 240 nm. Segura et al. [12] described a
quantitative analysis of diclofenac in plasma
using gas chromatography-mass spectrometry.
Sioufi et al. [13] determined diclofenac in
plasma and urine by using capillary gas
chromatography-mass spectrometry (GCMS).
High performance thin layer chromatography (HPTLC) can be used for identification
and for control of batch-to-batch consistency
in the stability testing of drugs and for
purposes of control throughout the entire
manufacturing process of drugs, as well as
quality control of the finished product. It has
the advantages of being sensitive, selective,
rapid, accurate and reproducible.
The present paper reports the development and validation of a new high performance thin layer chromatography (HPTLC)
method for determination of diclofenac
sodium in Voltaren emulgel, Diclogel and
Dosanac emulsiongel.
2. M ATERIALS AND M ETHODS
2.1 Chemicals
Diclofenac sodium (Reference Standard
No T191067, 99.9 %, Sigma, Switzerland),
Ethyl acetate (BDH laboratory supplies,
England), Toluene (Lab-scan analytical science,
Ireland), Glacial acetic acid (Farmitalia Carlo
Erba, Italy), Methanol (Lab-scan analytical
sciences, Ireland). HPTLC precoated plates
silica gel 60 F254 2010 cm, layer thickness 0.2

Chiang Mai J. Sci. 2006; 33(1)

mm (Merck, Germany). All other chemicals

used were analytical grade.
2.2 Samples
Diclogel (batch no. 30131; Polipharm
Co., Ltd.), Voltaren Emulgel (batch no.
2B101; OLIC (Thailand) Limited) and
Dosanac Emulsiongel (batch no. 925058;
Siam Bheasach Co., Ltd.).
2.3 Instruments
- CAMAG Automatic developing
system (Camag Muttenz,Switzerland).
- CAMAG TLC Scanner and
Computer Aided Testing Software (CATS)
evaluation software, deuterium lamp, scanning
by absorbance at 282 nm, evaluation via peak
area.
- Centrifuge (SIGMA, Scientific,
Germany).
- UV viewing system (ChromatoVUE C 70G, USA).
- UV-visible spectrophotometer
(Chromato-VUE C 70G, USA).
2.4 Standard Stock Solution
A standard stock solution of diclofenac
sodium (1,000 g.mL -1) was prepared in
methanol. Working standard solutions in a
range of 5-80 g.mL-1 were prepared by
dilution from this stock solution.
2.5 Sample Preparation
One gram of sample was accurately
weighed into a 100 mL beaker and dissolved
in 10 mL methanol. Then the sample solution
was transferred into a 25 mL volumetric flask
and adjusted to volume with methanol. The
solution was vortexed for 30 s then centrifuged at 4,000 rpm for 30 min. The supernatant containing diclofenac sodium was taken
and filtered by membrane filter (90 mm Dia.
Whatman). The filtrated supernatant was
used for further analysis.
2.6 HPTLC Method for Determining
Diclofenac Sodium
A series of standard solutions containing

125

5-80 g.mL-1 of diclofenac and the sample

solutions were applied respectively, applied the
3 mm band width by Camag 100 L sample
syringe (Hamilton, Bonaduz, Switzerland) on
a precoated silica gel aluminium plate 60 F254
(2010 cm) with 200 m layer thickness (E.
Merck, Darmstadt, Germany). A linomat IV
was employed with a constant rate of 3
s.L-1 and the space between two bands was
5 mm. Development was performed in an
Automatic Multiple Development (AMD)
system using toluene : ethyl acetate : glacial
acetic acid (60:40:1, v/v/v) as mobile phase,
[previous investigation has done as described
in the Results and Discussion section 3.1].
Densitometric scanning was performed on
Camag TLC scanner III in the reflectanceabsorbance mode (Zig-Zag) at 282 nm for
all measurements and operated by CATS
software. Concentrations of the compound
of chromatogram (spot) on the plate were
determined from the intensity of diffusely
reflected light. The spot areas were plotted
against diclofenac sodium concentrations.
Then the content of diclofenac sodium in each
sample was calculated by reference to the
calibration curve.
3. R ESUL
TS AND D ISCUSSION
ESULTS
3.1 Selection of Mobile Phase
During the development of the HPTLC
method four different compositions of
mobile phase including methanol : ethyl
acetate, methanol : ethyl acetate : glacial acetic
acid, toluene : ethyl acetate and toluene : ethyl
acetate : glacial acetic acid were tested. It was
found that the mobile phases, methanol : ethyl
acetate and methanol : ethyl acetate : glacial
acetic acid provided bad constituent separation with tailing occurred. When using toluene
: ethyl acetate as mobile phase the spots of
standard and sample moved slowly with
tailing occurred but no separation occurred
when using ethyl acetate as mobile phase. A
mixture of toluene : ethyl acetate : glacial acetic
acid (60:40:1, v/v/v) was proved to be the
best because it gave good resolution for
diclofenac sodium with the Rf of 0.51.

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Chiang Mai J. Sci. 2006; 33(1)

3.2 Method Validation

The following parameters has been used
to validate the developed HPTLC method
for the estimation of diclofenac sodium in
pharmaceutical formulations.
3.3 Sensitivity and Linearity
The sensitivity of the assay was determined in terms of limit of detection (LOD),
limit of quantitation (LOQ), linearity range
and correlation coefficient. The detection limit
of the method was investigated by applying
various concentrations of standard diclofenac
sodium solution on the precoated HPTLC
plate. After development, the spot areas were
quantified by densitometry. Limit of detection
(LOD) and limit of quantitation (LOQ) were
calculated from the standard deviation (S.D.)
of densitometric response and slope of curve
(S) using the equation;
LOD = 3.3 (S.D. / S)
LOQ = 10 (S.D. / S)
After densitometric analysis of diclofenac
sodium at 282 nm, the lowest amount of drug,
which could be detected was found to be 1

Area under curve

60000
50000
40000
30000
20000
10000
0

g.mL-1 (3 mole.L-1, which is more sensitive

than the UV spectrophotometric method [5]
and the lowest amount of drug which could
quantified was found to be 5 g.mL-1. The
calibration curve of diclofenac sodium
standard was found to be linear in the range
of 5-80 g.mL-1 (Figure 2). The mean values
(S.D.) of correlation coefficient, slope and
intercept are shown in Table 1.
3.4 Accuracy
The accuracy of the proposed method
was verified by analyzing the aliquots of
sample solutions equivalent to diclofenac
sodium 2 g.mL -1 spiked with various
concentrations of standard diclofenac sodium
solutions 25, 30 and 35 g.mL-1 respectively,
using the proposed procedure. The recovery
of each spiked standard diclofenac sodium
was calculated. Results are presented in Table
2.
Regression equation; Y = 149.92C +
6808.10 (r 2 = 0.9994) where C is the
concentration in g.mL-1 and Y is the peak
area.

y = 565.23x + 6821.8
2

R = 0.9994

20

40

60

Concentration (g/mL)

80

100

Figure 2. Linear calibration curve of diclofenac sodium 5-80 g.mL-1.

Table 1. Linear regression data for calibration curves (n=7) of diclofenac sodium obtained
from HPTLC.

Parameter
Linearity range
r2 S.D.
Slope S.D.
Intercept S.D.

Value
5-80 g.mL-1
0.9993 (7.0010-5)
564.08 ( 30.08)
6838.63 ( 20.98)

Chiang Mai J. Sci. 2006; 33(1)

127

Table 2. Recovery study of diclofenac sodium by TLC densitometric, standard addition

technique for determination of diclofenac sodium (n = 7).

Pharmaceutical sample
(marketed formulation)
(2 g.mL-1)
Diclogel

Concentration of
addition standard
(g.mL-1)
25
30
35

Voltaren emulgel

25
30
35

Dosanac emulsiongel

25
30
35

3.5 Precision
The intra-day reproducibility was
evaluated by analyzing the sample repeatedly
and inter-day reproducibility was evaluated by
analyzing the sample of diclofenac sodium
over a period of three days. Good precision
were obtained with %CV of 2.13 and 2.46
for intra-day and inter-day, respectively.
3.6 Application
The propose method was applied to the
determination of diclofenac sodium in
pharmaceutical preparation (emulsion gel
form). A comparative determination in the

% Recovery
(S.D.)
106.24 0.32
108.04 1.98
98.36 2.49
120.32 0.36
108.34 4.50
108.58 0.94
99.54 3.21
101.11 4.83
103.18 5.74

%CV
0.29
1.82
2.54
0.30
4.26
0.82
3.37
4.78
5.89

same samples were also investigated by

spectrophotometric method [5]. The results
are summarized in Table 3. Excellent correlation between the two methods was obtained.
The results were obtained from the proposed
method compared with those obtained by
using the spectrophotometric method with the
student t-test. The calculated t-value in all cases
were less than the tabulated value (2.44 for 6
degree of freedom at 95% confidence level).
These results indicate that the proposed
method does not differ significantly from
known method.

Table 3. Comparison between the HPTLC method and UV spectrophotometric

method for the determination of diclofenac sodium in dosage forms.

Pharmaceutical
Sample
Diclogel
Voltaren emulgel
Dosanac emulsiongel

% Labelled amount of diclofenac sodium

found (S.D.) *
HPTLC
UV spectrophotometric
method a
method a
94.61 0.06
96.26 0.98
97.87 0.11
96.78 0.48
94.81 0.03
95.31 0.91

*S.D. = standard deviation

Each value is the average of 7 determinations,
b
Tabulated t-value for p = 0.05 and six degrees of freedom is 2.44
a

Calculated
t-value b
1.47
1.30
0.47

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4. C ONCLUSION
The HPTLC method for determining
diclofenac sodium in pharmaceutical formulations was developed using toluene : ethyl
acetate : glacial acetic acid (60:40:1, v/v/v) as
mobile phase. The peak areas of the densitogram were quantified by densitometer at 282
nm. The limit of detection and limit of
quantitation were found to be 1 and 5
g.mL-1, respectively. The calibration curve
was linear over the range of 5-80 g.mL-1.
The mean values (S.D.) of correlation
coefficient, slope and intercept were found
to be 0.9993 (7.010-5), 564.08 (30.08) and
6838.63 (20.98), respectively. The method
was applied to the determination of diclofenac sodium in Diclogel, Voltaren emulgel
and Dosanac emulsiongel with the average
percentage recovery of 104.211.59,
112.411.93 and 101.274.59, respectively
and the % labelled amount of 94.610.06,
97.870.11 and 94.810.03, respectively. The
proposed method is simple, sensitive and
accurate with good precision is suitable for
routine analysis of this drug in formulations.
A CKNOWLEDGEMENT
We would like to thanks Graduated
School, Chiang Mai University and Faculty of
Pharmacy, Chiang Mai University for financial
and chemical supports.
R EFERENCES
[1] The United State Pharmacopeia USP 28,
2005, United States Pharmacopeial
Convention, INC. Asian Edition.
[2] The United State Pharmacopeia USP 25,
2002, United States Pharmacopeial
Convention, INC. Asian Edition.
[3] Martindale, The Extrapharmacopeia, The
Pharmaceutical Press, London, 29th
edition, 1980; 12.
[4] The British Pharmacopeia, 1998; 462463.
[5] Bucci R., Magri A.D., and Magri A.L.,
Determination of Diclofenac Salts in
Pharmaceutical Formulations, J. Anal.
Chem., 1998; 577-582.

Chiang Mai J. Sci. 2006; 33(1)

[6] Sena M.M., Chaudhry Z.F., Collins C.H.,

and Poppi R.J., Direct Determination of
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